The leukocyte common antigen (CD45) on human pre-B
leukemia cells: variant glycoprotein form expression during the cell exposure
to phorbol ester is blocked by a nonselective protein kinase inhibitor
H7

The human pre-B acute lymphoblastic leukemia cell line REH6 was utilized
for characterization of CD45 glycoprotein by monoclonal antibodies (mAb)
recognizing four distinct CD45 antigen specificities, i.e. nonrestricted
CD45, restricted CD45RA, CD45RB and CD45R0. Immunoprecipitation revealed
two antigen specificities on REH6 cells of m.w. 220 kDa and 190 kDa, both
presenting wide range of isoelectric point pI~6.0-7.5. Nonrestricted CD45
epitopes were not affected by the sialyl acid cleavage with sodium metaperiodate
or neuraminidase, but were sensitive to both, tunicamycin, the N-glycosylation
inhibitor and monensin, an inhibitor of protein transport through the Golgi
compartment. O-sialoglycoprotein endopeptidase from Pasteurella haemolytica
A1 partially cleaved CD45RA and CD45RB epitopes, while nonrestricted CD45
determinants were not affected by this enzyme. Limited proteolysis of this
antigen resulted in the appearance of 160-180 kDa peptide domains which
retained CD45 epitopes. Further, the treatment of cells with phorbol myristate
acetate (PMA) induced marked down-regulation of 220 and 190 kDa isoforms
and the appearance of new 210, 180 and 170 kDa variant glycoprotein forms
which were not found on parental cells. This PMA effect was not accompanied
by the programmed cell death and was markedly blocked by a nonselective
protein kinase (PK) inhibitor isoquinoline sulfonamide H7. Modulation of
CD45 by phorbol esters might serve as an in vitro model for an additional
insight into the function of CD45 in hematopoietic cells.

Hematologic effects of granulocyte colony-stimulating factor (G-CSF)
and erythropoietin (EPO) combination after priming intensive chemotherapy
in the treatment of female breast carcinoma are presented. In a previous
group treated with G-CSF alone, 36% of patients became anemic and had to
be transfused for correction of their anemia.
To the present study 11 consecutive patients with different stages of breast
carcinoma were admitted. All were given priming intensive chemotherapy
(epirubicin 150 mg/m2 and cyclophosphamide 1300 mg/m2)
followed by subcutaneous application of G-CSF at a dose of 5 µg/kg/day
and EPO 250 IU/kg/day. In cases where leucocyte counts dropped below 1
x 109/l and hemoglobin levels fell to 85 g/l administration
of growth factors was started. The therapy was stopped when normal leucocyte
count reached 4 x 109/l for G-CSF and hemoglobin level rose
to 115 g/l for EPO.
Our results show significant difference between MNC/Þl (min.), CD34+
cells/µl (min.), CFU-GM/ml (min.), BFU-E/ml (min.) and MNC/µl
(max.), CD34+cells/µl (max.), CFU-GM/ml (max.), BFU-E/ml
(max.) p < 0.01, with mean peak values of 16.9-fold for circulating
MNC/µl, 7.8-fold for CD34+ cells/µl, 23.4-fold for
CFU-GM/ml and 28.7-fold increase for BFU-E/ml. Side effects were minimal,
no infectious complications occurred, body temperature did not rise over
38°C and no corrections of anemia were needed.
It is concluded that the administration of G-CSF plus EPO combination following
intensive chemotherapy reduces hematologic toxicity and induces large amount
of hemopoietic progenitors suitable for autologous transplantation in women
with breast carcinoma.

We investigated activities of the cysteine protease cathepsin B (CB;
EC 3.4.22.1), the levels of reduced glutathione (GSH) and cysteine and
the activity of gamma-glutamyltransferase (gamma-GT; EC 2.3.2.2.) in squamous-cell
lung carcinoma (SQCLC) and the lung parenchyma specimens from surgically
treated patients. The basal CB activity, assayed in tissue extracts in
the absence of exogenous activators, was significantly higher in SQCLC
compared to the lung. The residual CB activity, remaining in tissue extracts
after preincubation at 37°C, was not any longer significantly different
in SQCLC and the lungs. The inhibited CB activity, calculated as the difference
between the basal and residual CB activities, was significantly higher
in SQCLC compared to the lung. In the case of the cysteine protease cathepsin
C (CC; EC 3.4.14.1), neither the basal nor the residual nor the inhibited
CC activities in SQCLC and the lung were significantly different. Compared
to CC, the powerfulness of endogenous cysteine protease inhibitors to inhibit
CB was much higher in both SQCLC and the lung. The cysteine protease inhibitors
from SQCLC and the lung which effectively inhibited CB could be related
to the inhibitors with an apparent Mr ranging from 10 000 to
30 000. Isoelectric focusing studies indicated significant differences
in the progress of inhibition of the activity of CB isoforms in SQCLC and
lung parenchyma extracts. The levels of both GSH and Cys were significantly
higher in SQCLC compared to the lung and the level of GSH was significantly
higher in Stage III tumors compared to Stage I tumors. The activity of
gamma-GT was not significantly different in SQCLC and the lung but it was
significantly higher in Stage I tumors compared to Stage III tumors and
showed a significant negative correlation with GSH level in SQCLC. Dithiothreitol
did not increase the basal activity of CB from SQCLC and the lung which
indicates that reversibly oxidized forms of CB do not accumulate in the
tumors and the lungs. The basal activity of CB from SQCLC and the lung
was competitively inhibited by Cys. Moreover, increasing Cys concentrations
had a modulatory effect on the basal activity of CB from SQCLC and the
lung which was featured by Cys-induced inhibition of CB activity and by
subsequent Cys-effected recovery of CB activity from its previous inhibition
by Cys.

Antiproliferation activity and the mechanism of
action of 9-bromo-5-morpholino-tetrazolo[1,5-c]quinazoline - potentional
anticancer drug

S. Jantová, ¼. Ružeková, Š. Stankovský,
K. Špirková

Department of Biochemistry and Microbiology, Faculty of Chemical
Technology, Slovak University of Technology, 812 37 Bratislava, Slovakia;
Department of Mutagenesis and Chemical Carcinogenesis, Cancer Research
Institute of the Slovak Academy of Sciences, Bratislava, Slovakia;
Department of Organic Chemistry, Faculty of Chemical Technology, Slovak
University of Technology, Bratislava, Slovakia

9-Bromo-5-morpholino-tetrazolo[1,5-c]quinazoline (BMTQ) at the two highest
tested concentrations (74.6; 29.8 µmol//l) induced retarded cytotoxic
effect. After 24 hours of culturing 23.1-98.8% of the cell population proliferated
but after 48 and 72 hours 6.4-80.4% of the cell population degenerated.
Other concentrations induced toxicity that was concentration- and time-dependent.
The cytolytic concentrations of BMTQ induced integrity damage of cytoplasmatic
membrane. The inhibition of cell cycle and the elevated content of proteins
in the cell exposed to the cytotoxic concentrations of BMTQ suggest that
the cells synthesize protein without entering into mitosis and that dying
cells are in the S-phase before death. BMTQ induced 1.75-3.01-times increase
of the level of ssDNA in comparison with the control.

We describe an alternative assay to determine genotoxicity. Its main
feature is that it combines two measures in a single experiment; the inhibition
of replicative DNA synthesis together with the stimulation of DNA repair.
We show that, in tests of four different genotoxic agents, the assay gives
results that are entirely consistent with what is known about the mode
of action of these agents. In addition, we have demonstrated that chemical
carcinogens requiring metabolic activation can be examined using a standard
procedure of incubation with a microsomal activating fraction. We consider
the combined assay for DNA synthesis inhibition and repair synthesis to
be a useful way for the rapid prescreening of chemicals suspected of genotoxic
activity on the level of mammalian cells.

The influence of melatonin on metabolic changes
in female rats induced by continuous irradiation and/or administration
of 7,12-dimethylbenz/a/anthracene

Metabolic profile is an important biological marker of neoplastic processes
not only in the tumor itself but also in the host organism. The neurohormone
melatonin has been implicated in experiments as an oncostatic agent.
Female Wistar:Han SPF rats (Velaz, Prague, Czech Republic) were irradiated
continuously for 15 days using a daily gamma rays dose of 96 mGy. At the
end of exposure one group of rats was administered 5 mg/kg b.w. of dimethylbenz/a/anthracene
(DMBA) intragastrically. During the period of exposure to ionizing radiation
a part of the animals was supplied with melatonin (M) at a concentration
of 20 µl/ml in drinking water. Selected parameters of lipid and carbohydrate
metabolisms and levels of selected hormones were determined 2, 30 and 100
days post-irradiation. The irradiation itself caused only small changes
in tissue lipids. The application of a single low dose (subthreshold from
the point of view of induction of mammary tumors) of DMBA caused more pronounced
changes in nonirradiated animals; of the changes observed an increase in
lipids in the liver, triacylglycerols (TG) in the thymus and decrease in
myocardial glycogen predominated. The intake (by drinking) of exogenous
M prevented the biochemical pattern of fatty liver in animals administered
DMBA in both groups, irradiated and nonirradiated. A prolonged effect of
exogenous M, demonstrated by prevention of increase in TG in the thymus
and of irradiated animals caused by administration of DMBA, was observed.
The mechanism of metabolic effect of M is not known. Additional experiments
are needed to explain the relationship between the beneficial effect of
M on metabolic changes and its presumable oncostatic effect in rats.

Family history of colorectal cancer is recognized as a risk factor for
the disease and the development of colorectal cancer represents a suitable
model for illustrating multistep tumor development. Bleomycin induced chromosome
sensitivity studies were done in 7 colorectal cancer families consisting
of 12 colorectal cancer patients and their 34 first degree relatives and
12 sporadic colorectal patients for comparison and identification of high
risk family members with genetic instability. All patients and 4 unaffected
relatives showed increased bleomycin sensitivity, which might be due to
defective DNA repair system. These four relatives may be classified as
high risk (without cancer at present) individuals. The study is being continued
in more number of familial colorectal cancer patients and their relatives
to arrive at definite conclusions.

ABVD chemotherapy of Hodgkin's disease

Z. Molnár, T. Schneider, E. Várady, T. Fleischmann

National Institute of Oncology, 1122 Budapest, Hungary

The authors report on their results in ABVD therapy, which was given
by 91 patients with Hodgkin's disease as first-line treatment. 78 patients
(85%) achieved complete, 10 (11%) partial remission, 3 (4%) did not respond
to therapy. In the follow-up period (36-223 months) 6 patients (7%) died
because of the progression of Hodgkin's disease. Serious side effects or
treatment-related death did not occur. Based on their results of ABVD,
the authors find ABVD chemotherapy effective as first-line treatment of
Hodgkin's disease.

Nuclear parameters were assessed by computer-assisted image analysis
in the the cells of abnormal epithelial formations in the acquired cystic
kidneys of two dialysis patients, the proximal and distal tubules of a
normal kidney and two well differentiated renal cell carcinomas. One acquired
cystic kidney contained many small clear celled foci and an 0.9 cm-size
clear celled lesion and the second one a papillary microadenoma. The clear
celled lesion was cytologically indistinguishable from the carcinomas.
The histomorphometrically gauged nuclear parameters were maximal and minimal
ferret diameters, averaged ferret diameter, aspect ratio, shape factor,
area, volume and specific length and width. Statistical evaluation evidenced
that the nuclear area, volume, aspect ratio and shape factor allowed for
the distinction between benign and malignant epithelial structures. The
medians of the nuclear parameters of atrophic tubules, cysts, clear celled
foci, papillary adenoma and clear celled lesion in the two acquired cystic
kidneys deviation from those of normal renal tubules and, in increasing
order of disparity, approached those of the carcinomas.

Mass scale cervical cytology which is the most accepted strategy for
the control of cervical cancer cannot be undertaken in developing countries
in view of paucity of resources, hence a need arises to examine alternate
strategy. The present exercise attempts to study the reduction in cumulative
incidence rate of cervical cancer by one life time selective screening.
The results revealed that cumulative incidence rate (CIR) of cervical cancer
per 100 000 in cohort of women during the age of 20 to 64 years was found
to be 2555.0 in the absence of screening. One life time selective screening
at the age of 40 and 45 years showed the reduction of 11.6 and 17.2% in
CIR respectively where as respective estimates in case of complete screening
at mentioned age groups were found to be 21.5% and 25%. In order to further
conserve the resources the strategy seems to be optimum for developing
countries.